scholarly journals CXCL13 shapes immunoactive tumor microenvironment and enhances the efficacy of PD-1 checkpoint blockade in high-grade serous ovarian cancer

2021 ◽  
Vol 9 (1) ◽  
pp. e001136
Author(s):  
Moran Yang ◽  
Jiaqi Lu ◽  
Guodong Zhang ◽  
Yiying Wang ◽  
Mengdi He ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
T Cells ◽  
B Cells ◽  
Tumor Microenvironment ◽  
Immune Checkpoint ◽  
Ex Vivo ◽  
Immune Checkpoint Blockade ◽  
Checkpoint Blockade ◽  
High Grade ◽  
Serous Ovarian Cancer

BackgroundMost patients with high-grade serous ovarian cancer (HGSC) lack an effective response to immune checkpoint blockade, highlighting the need for more knowledge about what is required for successful treatment. As follicular cytotoxic CXCR5+CD8+ T cells are maintained by reinvigoration by immune checkpoint blockade in tumors, we attempted to reveal the relationship between CXCR5+CD8+ T cells and the tumor microenvironment to predict immunotherapy responses in HGSC.Methods264 patients with HGSC from two cohorts and 340 HGSC cases from The Cancer Genome Atlas cohort were enrolled. Ex vivo and in vivo studies were conducted with human HGSC tumors and murine tumor models. The spatial correlation between CXC-chemokine ligand 13 (CXCL13), CXCR5, CD8, and CD20 was evaluated by immunohistochemistry and immunofluorescence. Survival was compared between different subsets of patients using Kaplan-Meier analysis. The therapeutic effect of CXCL13 and programmed cell death-1 (PD-1) blockade was validated using human HGSC tumors and murine models.ResultsHigh CXCL13 expression was associated with prolonged survival. Tumors with high CXCL13 expression exhibited increased infiltration of activated and CXCR5-expressing CD8+ T cells. Incubation with CXCL13 facilitated expansion and activation of CXCR5+CD8+ T cells ex vivo. CXCR5+CD8+ T cells appeared in closer proximity to CXCL13 in tumors and chemotaxis towards CXCL13 in vitro. The combination of CXCL13, CXCR5, and CD8+ T cells was an independent predictor for survival. In addition, CXCL13 was associated with clusters of CD20+ B cells. CD20+ B cells predicted better patient survival in the presence of CXCL13. Histological evaluation highlighted colocalization of CXCL13 with tertiary lymphoid structures (TLSs). TLSs carried prognostic benefit only in the presence of CXCL13. CXCL13 in combination with anti-PD-1 therapy retarded tumor growth in a CD8+ T-cell-dependent manner, resulting in increased infiltration of cytotoxic CD8+ T cells and CXCR5+CD8+ T cells.ConclusionsThese data define a critical role of CXCL13 in shaping antitumor microenvironment by facilitating the maintenance of CXCR5+CD8+ T cells in TLSs and support a clinical investigation for a combination of CXCL13 and PD-1 blockade therapy in HGSC.

scholarly journals Enhanced efficacy of simultaneous PD-1 and PD-L1 immune checkpoint blockade in high grade serous ovarian cancer

2020 ◽  
pp. canres.1674.2020
Author(s):  
Changxin Wan ◽  
Matthew P. Keany ◽  
Han Dong ◽  
Linah F. Al-Alem ◽  
Unnati M. Pandya ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Immune Checkpoint ◽  
Immune Checkpoint Blockade ◽  
Checkpoint Blockade ◽  
High Grade ◽  
Serous Ovarian Cancer

scholarly journals HDAC6 Inhibition Alleviates CLL-Induced T-Cell Dysfunction and Enhances Immune Checkpoint Blockade Efficacy in the Eμ-TCL1 Model

2020 ◽  
Vol 11 ◽  
Author(s):  
Kamira Maharaj ◽  
John J. Powers ◽  
Melanie Mediavilla-Varela ◽  
Alex Achille ◽  
Wael Gamal ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Immune Checkpoint ◽  
Immune Checkpoint Blockade ◽  
Tumor Burden ◽  
Checkpoint Blockade ◽  
Cell Phenotype ◽  
Cell Dysfunction ◽  
T Cell Dysfunction

Development of chronic lymphocytic leukemia (CLL) is associated with severe immune dysfunction. T-cell exhaustion, immune checkpoint upregulation, and increase of regulatory T cells contribute to an immunosuppressive tumor microenvironment. As a result, CLL patients are severely susceptible to infectious complications that increase morbidity and mortality. CLL B-cell survival is highly dependent upon interaction with the supportive tumor microenvironment. It has been postulated that the reversal of T-cell dysfunction in CLL may be beneficial to reduce tumor burden. Previous studies have also highlighted roles for histone deacetylase 6 (HDAC6) in regulation of immune cell phenotype and function. Here, we report for the first time that HDAC6 inhibition exerts beneficial immunomodulatory effects on CLL B cells and alleviates CLL-induced immunosuppression of CLL T cells. In the Eμ-TCL1 adoptive transfer murine model, genetic silencing or inhibition of HDAC6 reduced surface expression of programmed death-ligand 1 (PD-L1) on CLL B cells and lowered interleukin-10 (IL-10) levels. This occurred concurrently with a bolstered T-cell phenotype, demonstrated by alteration of coinhibitory molecules and activation status. Analysis of mice with similar tumor burden indicated that the majority of T-cell changes elicited by silencing or inhibition of HDAC6 in vivo are likely secondary to decrease of tumor burden and immunomodulation of CLL B cells. The data reported here suggest that CLL B cell phenotype may be altered by HDAC6-mediated hyperacetylation of the chaperone heat shock protein 90 (HSP90) and subsequent inhibition of the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway. Based on the beneficial immunomodulatory activity of HDAC6 inhibition, we rationalized that HDAC6 inhibitors could enhance immune checkpoint blockade in CLL. Conclusively, combination treatment with ACY738 augmented the antitumor efficacy of anti-PD-1 and anti-PD-L1 monoclonal antibodies in the Eμ-TCL1 adoptive transfer murine model. These combinatorial antitumor effects coincided with an increased cytotoxic CD8+ T-cell phenotype. Taken together, these data highlight a role for HDAC inhibitors in combination with immunotherapy and provides the rationale to investigate HDAC6 inhibition together with immune checkpoint blockade for treatment of CLL patients.

The Role of CXCL12-Mediated Aberrant Methylation and Tumor Microenvironment Remodeling in Carcinogenesis and Prognosis of Bladder Cancer

2021 ◽  
Author(s):  
Yiheng Du ◽  
Jin Cao ◽  
Xiang Jiang ◽  
Xiaowei Cai ◽  
Bo Wang ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
T Cells ◽  
Tumor Microenvironment ◽  
Immune Checkpoint ◽  
Bioinformatics Analysis ◽  
Immune Checkpoint Blockade ◽  
Checkpoint Blockade ◽  
Specific Marker ◽  
Cxcl12 Expression ◽  
Potential Association

Abstract Background Bladder cancer (BLCA) is the most common genitourinary tumor but lacks specific diagnostic biomarkers. Recent years have seen significant advances in the use and approval of immune checkpoint blockade (ICB) therapy to manage bladder cancer at advanced stages when platinum-based therapy has failed. The tumor microenvironment (TME) in bladder cancer is an essential player in patient's responsiveness to ICB therapy. Therefore, this manuscript explored the TME and identified CXCL12, a specific marker for inflammatory cancer associated fibroblasts(iCAFs), as potential molecular markers and therapeutic targets for bladder cancer. Methods We examined the gene expression profiles in the TCGA and GEO datasets to reveal the potential association of CXCL12 with the carcinogenesis and prognosis of bladder cancer. Methylation analysis of CXCL12 was performed using the UALCAN and MethSurv databases. The MCP-COUNTER, ESTIMATE, and TIDE algorithms were applied to estimate the TME components and predict immunotherapy responsiveness. An iCAFs signature was constructed using the ssGSEA algorithm. Bioinformatics analysis results were validated through immunohistochemistry of clinical samples. IMvigor210 cohort was used to validate bioinformatic predictions of therapeutic responsiveness to immune checkpoint inhibitors Results Our analysis revealed the potential association between aberrant promoter methylation of CXCL12 and bladder cancer carcinogenesis. CpG sites methylation of the CXCL12 gene body was associated with bladder cancer prognosis. Moreover, the expression level of CXCL12 exhibited a significant correlation with patients' pathological features and prognosis. Through gene enrichment analysis, CXCL12 was demonstrated to be associated with immune modulation and tumor microenvironment remodeling. The MCP-COUNTER and ESTIMATE algorithms verified significant correlations between CXCL12 and TME components, particularly CAFs, macrophages, and T cells. The TIDE algorithm provided evidence that T-cell clearance and dysfunction were more pronounced in bladder cancers characterized by high CXCL12 expression and high iCAFs scores, contributing to inferior responsiveness to ICB therapy. Patients who expressed high CXCL12 levels and had high iCAFs scores were likely to have less frequent FGFR3 mutation and a stromal-rich molecular subtype. Immunohistochemistry revealed that the close association of CXCL12 with iCAFs in bladder cancer potentially influenced the intratumoral infiltration of CD8 + T cells. CXCL12 expression in MIBC was increased significantly in NMIBC, which supports the bioinformatics analysis results. The IMvigor210 cohort confirmed the iCAFs score to be significantly associated with the responsiveness to immune checkpoint blockade therapy. Conclusions This work explores carcinogenesis and cancer-promoting roles of CXCL12 in bladder cancer. As a specific marker gene of iCAFs, CXCL12 potentially promotes bladder cancer progression by regulating the tumor microenvironment. Further exploration of the association between CXCL12 and iCAFs may unravel potential therapeutic targets for bladder precision medicine and improve the responsiveness of immune checkpoint blockade therapy.

scholarly journals Combined PD-L1 and TIM-3 blockade improves the expansion of fit human CD8+ antigen-specific T cells for adoptive immunotherapy

2021 ◽  
Author(s):  
Shirin Lak ◽  
Valérie Janelle ◽  
Anissa Djedid ◽  
Gabrielle Boudreau ◽  
Ann Brasey ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Immune Checkpoint ◽  
Cell Expansion ◽  
Ex Vivo ◽  
Immune Checkpoint Blockade ◽  
Checkpoint Blockade ◽  
Immune Checkpoints ◽  
T Cell Expansion ◽  
And Function

AbstractBackgroundThe stimulation and expansion of antigen-specific T cells ex vivo enables the targeting of a multitude of cancer antigens. However, clinical scale T-cell expansion from rare precursors requires repeated stimulations ex vivo leading to T-cell terminal effector differentiation and exhaustion that adversely impact therapeutic potential. We leveraged immune checkpoint blockade relevant to antigen-specific CD8+ human T cells to improve the expansion and function of T cells targeting clinically relevant antigens.MethodsA clinically-compliant protocol relying on peptide-pulsed monocyte-derived dendritic cells and cytokines was used to expand antigen-specific CD8+ targeting the oncogenic Epstein-Barr virus (EBV) and the tumor associated antigen (TAA) Wilms Tumor 1 (WT1) protein. The effects of antibody-mediated blockade of immune checkpoints applied to the cultures (T-cell expansion, phenotypes and function) were assessed at various time points. Genomic studies including single cell RNA (scRNA) sequencing and T-cell receptor sequencing were performed on EBV-specific T cells to inform about the impact of immune checkpoint blockade on the clonal distribution and gene expression of the expanded T cells.ResultsSeveral immune checkpoints were expressed early by ex vivo expanded antigen-specific CD8+ T cells, including PD-1 and TIM-3 with co-expression matching evidence of T-cell dysfunction as the cultures progressed. The introduction of anti-PD-L1 (expressed by the dendritic cells) and anti-TIM-3 antibodies in combination (but not individually) to the culture led to markedly improved antigen-specific T-cell expansion based on cell counts, fluorescent multimer staining and functional tests. This was not associated with evidence of T-cell dysfunction when compared to T cells expanded without immune checkpoint blockade. Genomics studies largely confirmed these results, showing that double blockade does not impart specific transcriptional programs or patterns on TCR repertoires. However, our data indicate that combined blockade may nonetheless alter gene expression in a minority of clonotypes and have donor-specific impacts.ConclusionsThe manufacturing of antigen-specific CD8+ T cells can be improved in terms of yield and functionality using blockade of TIM-3 and the PD-L1/PD-1 axis in combination. Overcoming the deleterious effects of multiple antigenic stimulations through PD-L1/TIM-3 blockade is a readily applicable approach for several adoptive-immunotherapy strategies.

Neoadjuvant Chemotherapy Modulates the Tumor Microenvironment in High-Grade Serous Ovarian Cancer

2021 ◽  
Author(s):  
Zhongling Zhuo ◽  
Min Tang ◽  
Hexin Li ◽  
Lili Zhang ◽  
Bingqing Han ◽  
...  
Keyword(s):  
Gene Expression ◽  
Ovarian Cancer ◽  
T Cells ◽  
Survival Analysis ◽  
Tumor Microenvironment ◽  
Neoadjuvant Chemotherapy ◽  
Mapk Pathway ◽  
Treatment Options ◽  
High Grade ◽  
Serous Ovarian Cancer

Abstract Background While surgical reduction with adjuvant chemotherapy is the traditional treatment for high-grade serous ovarian cancer (HGSOC), neoadjuvant chemotherapy (NACT) has increasingly been applied. This work aims to investigate the expression profiles before and after NACT, explore changes in the tumor microenvironment, expand current treatments, and design a combination of treatment options for patients. Methods We downloaded 326 pre-NACT RNA sequencing data and 37 matched pre- and post-NACT samples from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Differentially expressed genes (DEGs) were determined with EdgeR, and Gene Ontology analysis was performed to identify the clusters responsible for the biological processes and pathways of HGSOC. Immune infiltration was analyzed using Single-sample Gene Set Enrichment Analysis (ssGSEA) and CIBERSORT. Kaplan-Meier (KM) survival analysis was performed to assess prognosis, and the potential correlations between modules and phenotypes were explored using weighted gene co-expression network analysis (WGCNA). Results After NACT, a total of 352 genes showed significant changes in RNA expression, among which 180 genes were up-regulated and 172 down-regulated. The most influential pathway was the positive regulation of mitogen-activated protein kinase (MAPK) cascade. Correlation analysis and KM survival analysis showed that overexpression of MAPK cascade genes correlated with shorter survival time in HGSOC patients. ssGSEA results showed that the expressions of anti-tumor cells (central memory CD4+ T cell and central memory CD8+ T cell) and pro-tumor cells (neutrophil and dendritic cells) were significantly increased after NACT. CIBERSORT showed that the abundances of memory B cells, NK cells, and monocytes were increased and the abundance of plasma cells was decreased after NACT. WGCNA and KM survival analysis showed that a lower abundance of Regulatory T cells (Tregs) was correlated with a better prognosis. Conclusions Gene expression of the MAPK pathway is up-regulated and the abundance of CD4+ T regulation cell decreases after NACT. Thus, the MAPK pathway may promote the differentiation of CD4+ T cells into Th17 cells while inhibiting Tregs development. The inhibited Tregs' development can lead to a better prognosis. Therefore, it is speculated that Tregs inhibitors combined with platinum-based NACT are potential treatment options for HGSOC.

scholarly journals IMMU-27. IMMUNE CHECKPOINT BLOCKADE OF EX VIVO EXPANDED T CELLS FOR USE IN ADOPTIVE CELLULAR THERAPY (ACT) FOR GLIOBLASTOMA

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi124-vi125
Author(s):  
Elizabeth Ogando-Rivas ◽  
Changlin Yang ◽  
Paul Castillo ◽  
Anjelika Dechkovskaia ◽  
Duane Mitchell
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
Immune Checkpoint ◽  
Cellular Therapy ◽  
Ex Vivo ◽  
Immune Checkpoint Blockade ◽  
Checkpoint Blockade ◽  
Study Objective

Abstract BACKGROUND Despite aggressive treatments, GBM continues to have unacceptably high mortality rates. Immune-checkpoint blockade and ACT have shown excellent results in other solid tumors, especially in melanoma. Unfortunately, these results have not been extrapolated to GBM. We have developed a novel platform for ACT using tumor mRNA-pulsed dendritic cells(DCs) to in-vitro expand polyclonal populations of tumor-reactive T-cells. This platform has shown promising effects in preclinical brain tumor models (Flores et al OncoImmunology 2015, Wildes et al CCR 2018, Flores et al NatureComm 2018) and being evaluated in clinical trials at UF Health (NCT02465268,NCT03334305). STUDY OBJECTIVE Evaluate whether immune-checkpoint blockade during ex-vivo expansion of antigen-specific T-cells impact their use in ACT. METHODS CMVpp65 was used as model antigen for in-vitro activation of T-cells. Mature pp65 mRNA-pulsed DCs from CMV+ healthy donors were co-cultured with T-cells in IL2-containing medium for 15days. We tested four checkpoint inhibitor groups: PD1(n= 6), PDL1(n= 4), TIM3(n= 7) and PD1+TIM3(n= 6) that were compared with non-blockade group, respectively. Checkpoint blockade was performed every 3days. T-cell proliferation, immune-phenotyping, and IFN-g release were analyzed. RESULTS Cell proliferation was lower in all the blockade groups but significantly lower in the TIM3 (p= 0.03) and TIM3+PD1 (p= 0.01) blockade groups. TIM3 expression was significantly lower in the TIM3 (p= 0.006) and PD1+TIM3 blockade groups (p= 0.0001). There was a trend of reduced pp65 tetramer positive in the TIM3 and PD1+TIM3 blockade groups (PD1+TIM3 subgroup at 3mcg/mL, p= 0.02) and lower INFg release in the TIM3 and PD1+TIM3 blockade groups. CONCLUSION The exact role of checkpoints during expansion of T-cells for ACT is not well understood. In our study checkpoint blockade with PD-1 or TIM-3 alone or in combination did not enhance T-cell expansion or function, in fact, appeared to have an inhibitory effect on measured parameters. Our results suggest that TIM-3 may have an activating role in our system.

EXTH-42. COMBINATION OF MAPK PATHWAY INHIBITORS AND IMMUNE CHECKPOINT BLOCKADE IN BRAF-MUTANT HIGH-GRADE GLIOMA

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi172-vi173
Author(s):  
Jong-Whi Park ◽  
Stefan Grossauer ◽  
Wei Wang ◽  
Mathieu Daynac ◽  
Sharon Pitteri ◽  
...  
Keyword(s):  
T Cells ◽  
Immune Checkpoint ◽  
Mapk Pathway ◽  
High Grade Glioma ◽  
Immune Checkpoint Blockade ◽  
Checkpoint Blockade ◽  
Braf V600e ◽  
High Grade ◽  
Pathway Inhibition ◽  
Braf Mutant

Abstract BACKGROUND Despite successes, clinical MAPK pathway inhibitors show limited anti-tumor activity in the majority of patients with BRAF-mutant high-grade glioma. Because of the presence of higher fraction of CD8+ tumor-infiltrating T cells in MAPK pathway-altered glioma, we explored the possibility that combined BRAF and MEK inhibition with immune checkpoint blockade enhances anti-tumor response. METHODS We engineered mice to carry BRAF V600E expression and CDKN2A deletion in various hemispheric areas. We treated syngeneic tumor-bearing mice with dabrafenib, trametinib, anti-PD-L1 and anti-CTLA-4 antibodies, and analyzed the tumor immune infiltrate by high-dimensional single-cell mass cytometry (CyTOF). RNA sequencing and Gene Set Enrichment Analysis were conducted using patient-derived BRAF-mutant glioma lines upon the inhibitor treatment. RESULTS The transcriptome analysis demonstrated that antigen processing and presentation feature is strongly enriched upon dual MAPK pathway inhibition. Consistent with these molecular changes, dabrafenib and trametinib treatment led to dynamic changes in tumor-infiltrating immune cells, including CD8+ and CD4+ T cells. In line with this, combination of MAPK pathway and immune checkpoint inhibitors elicit a significant survival benefit over MAPK pathway inhibition alone in mice with orthotopic BRAF-mutant glioma. CONCLUSIONS Clinically relevant molecular targeted therapy by dabrafenib and trametinib and immune checkpoint blockade synergize in pre-clinical models.

scholarly journals Stromal HIF2 Regulates Immune Suppression in the Pancreatic Cancer Microenvironment

2021 ◽  
Author(s):  
Yanqing Huang ◽  
Carolina J. Garcia Garcia ◽  
Daniel Lin ◽  
Nicholas D. Nguyen ◽  
Tara N. Fujimoto ◽  
...  
Keyword(s):  
Tumor Microenvironment ◽  
Tumor Progression ◽  
Immune Checkpoint ◽  
Cellular Response ◽  
Ex Vivo ◽  
Smooth Muscle Actin ◽  
Immune Checkpoint Blockade ◽  
Checkpoint Blockade ◽  
Ductal Adenocarcinoma ◽  
Pancreatic Tumors

Background & Aims. Pancreatic ductal adenocarcinoma (PDAC) has a hypoxic, immunosuppressive stroma, which contributes to its resistance to immune checkpoint blockade therapies. The hypoxia-inducible factors (HIFs) mediate the cellular response to hypoxia, but their role within the PDAC tumor microenvironment remains unknown. Methods. We used a dual recombinase mouse model to delete Hif1α or Hif2α in α-smooth muscle actin (αSMA)-expressing cancer-associated fibroblasts (CAFs) arising within spontaneous pancreatic tumors. The effects of CAF-Hif2α expression on tumor progression and composition of the tumor microenvironment were evaluated by Kaplan-Meier analysis, quantitative real-time polymerase chain reaction, histology, immunostaining, and by both bulk and single-cell RNA sequencing. CAF-macrophage crosstalk was modeled ex vivo using conditioned media from CAFs after treatment with hypoxia and PT2399, a HIF2 inhibitor currently in clinical trials. Syngeneic flank and orthotopic PDAC models were used to assess whether HIF2 inhibition improves response to immune checkpoint blockade. Results. CAF-specific deletion of HIF2, but not HIF1, suppressed PDAC tumor progression and growth, and improved survival of mice by 50% (n = 21-23 mice/group, Log-rank P = 0.0009). Deletion of CAF-HIF2 modestly reduced tumor fibrosis and significantly decreased the intratumoral recruitment of immunosuppressive M2 macrophages and regulatory T cells. Treatment with the clinical HIF2 inhibitor PT2399 significantly reduced in vitro macrophage chemotaxis and M2 polarization, and improved tumor responses to immunotherapy in both syngeneic PDAC mouse models. Conclusions. Together, these data suggest that stromal HIF2 is an essential component of PDAC pathobiology and is a druggable therapeutic target that could relieve tumor microenvironment immunosuppression and enhance immune responses in this disease.

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